Fat Mass and Obesity Associated Gene Polymorphism and the Risk of Polycystic Ovary Syndrome: A Meta-analysis.

Background: We aimed to elucidate the association between fat mass and obesity associated gene (FTO) polymorphism and the risk of polycystic ovary syndrome (PCOS) by meta-analysis. Methods: We searched PubMed and Embase databases to find the relevant studies. Odds ratios (ORs) and their corresponding 95% confidence intervals (CIs) were used for pooled analysis. Statistical analyses were carried out by using R 3.12 software. Heterogeneity was assessed using I 2 and Q statistics. I 2 >50% or P<0.05 was considered as heterogeneity statistically, and random effects model was used for pooled analysis. Otherwise, fixed-effect model was used. Results: Twelve eligible studies that published from 2008 to 2015 were included in this meta-analysis. The pooled analyses showed that rs9939609 polymorphism of FTO gene was significantly associated with risk of PCOS under A vs. T, AT vs. TT, AA vs. TT, AA vs. AT+TT and AA+AT vs. TT genetic models. However, for rs8050136 and rs1421085, significant association was only found under recessive genetic model. Conclusion: rs9939609 variation of FTO gene is significantly associated with risk of PCOS. However, the association between rs8050136, rs1421085, and PCOS is still unclear and needs further confirmation.


Introduction
Polycystic ovary syndrome (PCOS) is a common endocrine disease among women of reproductive age (1)(2)(3). The typical symptoms of PCOS include infertility, obesity, and hairiness, caused by elevated male hormone. PCOS can also arouse anxiety and depression of the patients (4). Up to now, the treatment of PCOS is still a challenge for physicians and medical researchers. The treatments are mainly adopted depending on the symptoms of the PCOS patients. For examples, letrozole and clomiphene are used to treat the infertility of PCOS (5). Some insulin-sensitizing drugs such as metformin, usually used to manage type 2 diabetes mellitus, are also applied to treat PCOS now (6). However, individual variations of the PCOS should be considered. For instance, a special PCOS patient may have normal male hormone levels. In addition, PCOS is associated with hyperinsulinemia and peripheral insulin resistance, and obesity can aggravate both abnormalities (7). Because of the difficult treatment of PCOS, the medical researchers have tried to consider the risk factors associated with PCOS to prevent prevalence of the disease. Genetic and environmental factors have been identified to be associated with PCOS. The fat mass and obesityassociated gene (FTO) as an obesity candidate gene has been related to PCOS susceptibility (8). However, the results of the studies were confused. For example, a previous study (9) states that FTO is significantly associated with the risk of PCOS, but another study (10) reports no association between the two entities.
In this study we systematically meta-analyzed the genotypes of FTO gene and risk of PCOS, thus, to identify key variations of FTO gene associated with risk of PCOS.

Source of data
We searched two English databases PubMed and Embase using the search terms of (PCOS OR polycystic ovarian syndrome OR Polycystic ovary syndrome) and (FTO OR fat mass and obesity associated gene OR s1121980 OR rs1421085 OR rs1558902 OR rs8050136 or rs17817449 or rs9939609 or rs9930506). The deadline of the search was Mar 2016. Web of science and Google scholar were also used for checking leakage.

Inclusion and exclusion criteria
Inclusion criteria included: 1) the study reported the association between FTO and FCOS; 2) the study provided the data for calculating the distribution of FTO in PCOS patients and non-PCOS population. The reviews, reports, comments and letters were excluded from this analysis.

Data extraction and quality assessment
The following data extracted from included studies: name of the first author, publication year, location of the study, the number of PCOS patients (cases) and non-PCOS population (controls), age of the participants and Body Mass Index (BMI). The number of each genotype in cases and controls was also extracted. Quality assessment was conducted using Newcastle-Ottawa Scale (NOS) (11) independently. The disagreements in the processes of data extraction and quality assessment were resolved by discussing with Yongxia Chen.

Statistical analysis
Hardy-Weinberg equilibrium (HWE) test was performed using chi-square (12). Pooled analysis was carried out using R 3.12 software with the effect size of odds ratios (ORs) and their corres-ponding 95% confidence interval (CI). Heterogeneity was assessed using I 2 and Q statistics (13). I 2 >50% or P<0.05 was considered as heterogeneity statistically, and random effects model was used for pooled analysis. Otherwise, a fixedeffect model was used. Publication bias was assessed by using Egger's test and P<0.05 represented a significant publication bias (14).

Characteristics of included studies
The characteristics of 12 case-control studies were shown in Table 1. The publication year of included studies ranged from 2008 to 2015. These studies distributed in Korea, America, Brazil, and China. The major genotypes of FTO gene were rs17817449, rs1121980, rs1558902, rs11642841, rs17817449, rs9939609, rs8050136 and rs1421085. The quality assessment showed relative high quality with 6-8 scores of these in-cluded studies. HWE analysis ( Table 2) showed that all the controls at rs9939609 loci conformed to HWE. However, two populations (17,22) at rs8050136 loci and one population (15) at rs1421085 deviated from HWE (P<0.05).

Pooled analysis
We meta-analyzed the association between PCOS and FTO gene polymorphisms at loci of rs9939609 (A vs T), rs8050136 (A vs C) and rs1421085 (C vs T) under allele, additive, recessive and dominant genetic models. As shown in Fig. 2, heterogeneity test showed significant heterogeneity in rs9939609 under allele (A vs. T, I 2 =61.2%), additive (AT vs. TT, I 2 =61.2%), recessive (AA vs. AT+TT, I 2 =67.6%) and dominant genetic models (AA+AT vs. TT, I 2 =62.5%). Therefore, the random effects model was used for pooling analysis. The analyses under other genetic models were pooled using fixed effect model.   Fig. 4), respectively.
Egger's test (Table 3) showed no publication bias in any genetic model of FTO gene (P>0.05), which indicated that the pooled results in this meta-analysis were credible.

Discussion
In this study, we conducted a meta-analysis on the association between FTO gene polymorphisms and the risk of PCOS. The mutation at rs9939609 loci of FTO could significantly increase the risk of PCOS. However, no reliable evidence can identify the relationship between the other two SNPs of FTO (rs8050136 and rs1421085) and PCOS. Our results are opposite to a previous metaanalysis which states that rs9939609 variant of FTO gene has no association with PCOS (24). Rs9939609 is a common polymorphism of FTO gene and has been identified to link with obesity. PCOS patients had significantly higher mean BMI compared to the people without PCOS (25). However, most of the PCOS patients (81.3%) were not obese. Interestingly, the researchers (25) found that rs9939609 was not significantly associated with PCOS, but significantly associated with obese PCOS. Therefore, rs9939609 variant was not the major decisive factor of PCOS, and FTO might influence PCOS through an association with obesity. However, rs9939609 in FTO was associated with the risk of Chinese PCOS women both in obese and lean cases (19). The same result was found in rs1421085 and rs8050136 loci in Korean PCOS women after adjusting BMI (8). Thus, we guess that race might play a role in the association between FTO gene polymorphism and PCOS. Unfortunately, we did not adjust the obesity and race of the participants by meta-regression in this meta-analysis because of lack of the included studies and demographic characteristics of the included populations. This may be also a source of significant heterogeneity in pooled analysis. Nonetheless, it could remind us to consider these important factors when performing experiment design in future. Except for obesity and race, metabolic syndrome may be another important factor influencing the association between FTO polymorphism and PCOS. Rs1421085 (C/T) polymorphism in FTO showed a significant association with obese PCOS women or PCOS patients with metabolic syndrome, but not associated with lean PCOS patients or controls (15). In addition, the type of PCOS should be also considered. In the 12 included studies, only one referred to the type of PCOS (17), and no association was found between FTO variation and insulin resistant phenotype of PCOS in that study. Moreover, it is very necessary to adjust more potential confounders to clarify the association between FTO polymorphism and PCOS.
There are several limitations in this meta-analysis. Firstly, some covariates were not adjusted because of lack of studies and incomplete data. Secondly, the influences of some variates of FTO (rs1121980, rs1558902, rs11642841, and rs17817449) on PCOS were not meta-analyzed due to insufficient data. Thirdly, HWE test showed disequilibrium in controls in several included studies. Therefore, these controls may not predict the condition of the population in a certain region. Despite all this, our study can provide an important reference for the further researchers and help them understand the association between FTO polymorphism and PCOS systematically.

Conclusion
Rs9939609 SNP variation in FTO gene can significantly increase the risk of PCOS in women. However, rs8050136 and rs1421085 are associated with PCOS under recessive model, but no association under other genetic models. Therefore, the association between these two SNPs and PCOS are still unclear. Further studies with large-scale and high quality are needed to elucidate the association between them. Our study would provide reference for understanding the relationship between FTO and the risk of PCOS and help clinicians take strategy to treat PCOS.

Ethical considerations
Ethical issues (Including plagiarism, informed consent, misconduct, data fabrication and/or falsification, double publication and/or submission, redundancy, etc.) have been completely observed by the authors.